Project Summary Endothelial dysfunction induced by barrier dysfunction and hyper-coagulation is one of the major pathological features of sepsis for which no effective treatment exists. Therefore, identifying novel molecular mechanisms regulating the cross-talk of vascular barrier function and coagulation during sepsis is of great scientific and therapeutic interest. The current proposal arises from our discovery that orphan nuclear receptor Nur77 is a critical determinant controlling vascular inflammation, hyperpermeability, and microvascular thrombosis in response to septic injury. Our data show that Nur77 can simultaneously suppress NF-kB activation while exerting potent anti-thrombotic effects by increasing thrombomodulin (TM) expression and upregulating levels of activated protein C (APC) in vascular endothelial cells (ECs). The protective effects of Nur77 are dependent on early up-regulation in the vascular endothelium and that failure to increase its expression immediately after injury leads to a dramatic increase in vascular permeability and inflammation. Further, we show that the barrier protective effects of Nur77 appear to be mediated through upregulation of sphingosine kinase 1 (SphK1), as SphK1 and its downstream product sphingosine-1-shosphate (S1P), are substantially reduced in Nur77- deficient ECs exposed to lipopolysaccharide (LPS). Based on the collective strength of these data, we hypothesize that Nur77 serves as an essential brake to limit the severity of endothelial dysfunction in sepsis, and targeted activation of Nur77 can effectively treat sepsis by its concomitant anti-inflammatory, anti- hyperpermeability, and anti-thrombotic activities. To test this hypothesis, 3 specific aims are proposed. Aim 1 will establish the endothelial specific role of Nur77 in maintaining vascular homeostasis in sepsis by assessing whether mice with targeted deletion of Nur77 in endothelium are more susceptible to septic injury as a result of increased vascular leakage, inflammation, and prothrombotic activities. Aim 2 will delineate the molecular mechanisms by which Nur77 improves barrier function in endothelial cells. The regulation of the SphK1/S1P pathway by Nur77 and the functional importance of the Nur77/S1P axis in the Nur77-mediated stabilization of endothelial barrier function will be determined. Aim 3 will establish the therapeutic capacity of Nur77 activators in the treatment of sepsis. Our study will elucidate a critical mechanism underlying the regulation of endothelial barrier function and inflammation in sepsis and establish that this mechanism can be targeted to treat sepsis. The predicted utility and feasibility of such targeting in humans favors the translational potential and ultimate impact of the proposed studies.